Electrochemical sensing methods and apparatus for determining drug uptake and retention in cells

Abstract

This invention provides methods and apparatuses for the rapid assessment of cell permeability by a drug. More particularly, described herein a method of determining membrane permeability (influx and/or efflux) of a cell to a drug, the method including: (a) obtaining a biological sample; (b) dispersing at least one cell from the biological sample to a discrete location; (c) exposing the at least one cell to one member of a drug panel in a drug solution, wherein the drug panel is composed of drugs of a given concentration; (d) incubating the at least one cell from the biological sample in the drug for a given time; (e) obtaining at least one electro-analytical measurement of the discrete location adjacent the at least one cell.

What is claimed is: 
     
         1 . A method of determining membrane permeability of a cell to a drug, the method comprising: (a) obtaining a biological sample; (b) dispersing at least one cell from the biological sample to a discrete location or attached to a discrete substrate; (c) exposing the at least one cell to one member of a drug panel in a drug solution, wherein the drug panel is comprised of drugs of a given concentration; (d) incubating the at least one cell from the biological sample in the drug for a given time; (e) obtaining at least one electro-analytical measurement of the discrete location adjacent the at least one cell. 
     
     
         2 . The method of  claim 1 , wherein the method further comprises exchanging the drug solution for a drug-less solution. 
     
     
         3 . The method of  claim 2 , wherein the method further comprises incubating the at least one cell from the biological sample in the drug-less solution for a given time. 
     
     
         4 . The method of  claim 3 , wherein the method further comprises obtaining at least one further electro-analytical measurement of the discrete location adjacent to the at least one cell. 
     
     
         5 . The method of any one of  claims 1 - 4 , wherein the drug is an electro-active drug. 
     
     
         6 . The method of any one of  claims 1 - 4 , wherein the drug is selected from: an antibiotic drug and an anticancer drug. 
     
     
         7 . The method of  claim 6 , wherein the antibiotic drug is selected from one or more of the following: ampicillin; penicillin; amoxicillin; neomycin; tobramycin; ciprofloxacin; levofloxacin; norfloxacin; enrofloxacin; ofloxacin; linezolid; tetracycline; and azithromycin; or a hybrid of two or more antibiotic drugs. 
     
     
         8 . The method of  claim 6 , wherein the anticancer drug is selected from one or more of the following: 6-mercaptopurine; 5-fluorouracil; gemcitabine; doxorubicin; mitoxantrone; epirubicin; daunorubicin; valrubicin; cisplatin; temodal; oxaliplatin; carboplatin; etoposide; ifosfamide; erlotinib; irinotecan; and roscovitine; or a hybrid of two or more anti-cancer drugs. 
     
     
         9 . The method of any one of  claims 1 - 8 , wherein the drug panel is comprised of multiple drugs each at a variety of concentrations or combinations of drugs each at a variety of concentrations. 
     
     
         10 . The method of any one of  claims 1 - 9 , wherein the biological sample comprises bacteria isolated from a patient. 
     
     
         11 . The method of any one of  claims 1 - 9 , wherein the biological sample comprises a cancer biopsy from a patient. 
     
     
         12 . The method of any one of  claims 1 - 11 , wherein the electro-analytical measurement is made by one or more of the following: linear sweep voltammetry (LSV); cyclic voltammetry (CV); differential pulse voltammetry (DPV); differential pulse anodic stripping voltammetry (DPASV); square wave voltammetry (SWV); adsorptive stripping linear sweep voltammetry (AdSLSV); electrochemical impedance spectroscopy (EIS); chronoamperometry (CA); chronopotentiometry (CP); chronocoulometry (CC); impact chemistry (IC); scanning ion conductance microscopy (SICM); scanning electrochemical cell microscopy (SECCM); scanning photoelectrochemical microscopy (SPECM); and scanning electrochemical microscopy (SECM). 
     
     
         13 . The method of any one of  claims 1 - 12 , wherein electrode is optimized for the electro-active drug or electro-active drugs at the discrete location. 
     
     
         14 . An apparatus, the apparatus comprising (a) cell retention array having a plurality of array locations; and (b) a corresponding electrode array, wherein each electrode corresponds to each array location or a group of electrode locations and wherein the electrode is selected to be operable for a corresponding drug solution. 
     
     
         15 . A microfluidic device, the microfluidic device comprising (a) a plurality of cell retention locations; and (b) a corresponding electrode for each cell retention location or locations and wherein the electrodes are selected to be operable for a corresponding drug solution which might be delivered to the retention location or locations. 
     
     
         16 . The microfluidic device of  claim 15 , wherein the microfluidic device further comprises a system for fluid exchange at one or more of the retention locations.\ 
     
     
         17 . An apparatus, the apparatus comprising (a) a plurality of cell retention substrates; and (b) a corresponding electrode associated with each cell retention substrate, wherein the electrode is selected to be operable for a corresponding drug solution. 
     
     
         18 . The apparatus of  claim 17 , wherein the cell retention substrates are beads. 
     
     
         19 . An apparatus, the apparatus comprising:
 a cell retention array having a plurality of array locations;   wherein each cell retention array location corresponds to an electrode, and wherein each electrode is suitable for deposition of a cell on the surface of the electrode.   
     
     
         20 . The apparatus of  claim 19 , wherein each electrode is operable to retain the cell on the electrode by dropcasting. 
     
     
         21 . The apparatus of  claim 20 , wherein the each electrode is operable to receive a drug solution. 
     
     
         22 . A method of determining membrane permeability of a cell to a drug, the method comprising: (a) obtaining a biological sample; (b) dispersing at least one cell from the biological sample to a surface or attached to a surface; (c) exposing the at least one cell to a drug solution, wherein the drug solution has a given drug concentration; (d) incubating the at least one cell from the biological sample in the drug solution for a given time; (e) obtaining at least one electro-analytical measurement of the at least one cell by impact chemistry (IC), whereby the at least one cell from the biological sample is made to collide with an electrode. 
     
     
         23 . The method of  claim 22 , wherein the method further comprises exchanging the drug solution for a drug-less solution. 
     
     
         24 . The method of  claim 23 , wherein the method further comprises incubating the at least one cell from the biological sample in the drug-less solution for a given time. 
     
     
         25 . The method of  claim 24 , wherein the method further comprises obtaining at least one further electro-analytical measurement of the at least one cell from the biological sample. 
     
     
         26 . The method of any one of  claims 22 - 25 , wherein the drug is an electro-active drug. 
     
     
         27 . The method of any one of  claims 22 - 26 , wherein the drug is selected from: an antibiotic drug and an anticancer drug. 
     
     
         28 . The method of  claim 27 , wherein the antibiotic drug is selected from one or more of the following: ampicillin; penicillin; amoxicillin; neomycin; tobramycin; ciprofloxacin; levofloxacin; norfloxacin; enrofloxacin; ofloxacin; linezolid; tetracycline; and azithromycin; or a hybrid of two or more antibiotic drugs. 
     
     
         29 . The method of  claim 27 , wherein the anticancer drug is selected from one or more of the following: 6-mercaptopurine; 5-fluorouracil; gemcitabine; doxorubicin; mitoxantrone; epirubicin; daunorubicin; valrubicin; cisplatin; temodal; oxaliplatin; carboplatin; etoposide; ifosfamide; erlotinib; irinotecan; and roscovitine; or a hybrid of two or more anti-cancer drugs. 
     
     
         30 . The method of any one of  claims 22 - 29 , wherein the dispersing of at least one cell from the biological sample to the surface or attached to the surface drug panel repeated on multiple discrete surfaces so that multiple drugs at a variety of concentrations or combinations of drugs each at a variety of concentrations are available for IC electro-analytical measurement. 
     
     
         31 . The method of any one of  claims 22 - 30 , wherein the biological sample comprises bacteria isolated from a patient. 
     
     
         32 . The method of any one of  claims 22 - 30 , wherein the biological sample comprises a cancer biopsy from a patient. 
     
     
         33 . The method of any one of  claims 22 - 32 , IC electrode is a wire electrode. 
     
     
         34 . The method of any one of  claims 22 - 33 , wherein electrode is optimized for the electro-active drug or electro-active drugs at the discrete location.